Temporal dynamics of substance P and its association with cytokine release syndrome after CD19 CAR-T therapy in pediatric B-ALL

BACKGROUND: Cytokine Release Syndrome (CRS) remains a major toxicity associated with chimeric antigen receptor T (CAR-T) cell therapy, particularly in pediatric patients. Although neuroimmune mediators have been implicated in systemic inflammation, the role of neuropeptides such as Substance P (SP) in CRS has not yet been explored in this setting. METHODS: Plasma SP levels were measured using an enzyme-linked immunosorbent assay (ELISA) and analyzed longitudinally at predefined time points (day - 1, +7, + 14, and + 28) and during CRS when additional samples were available in 18 pediatric and young adult patients with relapsed/refractory B-cell acute lymphoblastic leukemia (R/R B-ALL) treated with CD19 CAR-T therapy (tisagenlecleucel) in a real-world clinical setting. SP dynamics were correlated with severity of CRS, inflammatory biomarkers, and CD19 CAR-T expansion kinetics. RESULTS: Baseline SP levels were similar between patients who developed severe CRS (grade ≥ 3) and those with non-severe CRS. In contrast, the increase in SP levels from baseline (ΔSP) was significantly higher in patients who developed severe CRS (1523 pg/mL vs. 189 pg/mL, p = 0.01). A peak in SP levels was observed at the onset of severe CRS in all three patients, coinciding with the beginning of clinical toxicity and occurring shortly before the elevation in ferritin levels. In these three cases, IL-6 levels increased in close temporal proximity to the rise in SP at the time of maximum CRS severity. Overall, SP levels declined by day + 14 post-infusion, shortly after the peak of CAR-T expansion. This decline coincided temporally with the administration of anti-CRS treatment in the subset of patients with severe CRS. SP levels later increased toward the end of the observation period (around day + 28), approaching baseline values. CONCLUSIONS: These findings suggest that SP dynamics may be linked to the early inflammatory response during CRS and support further investigation of the SP-NK1 receptor axis as a potential therapeutic target to modulate CD19 CAR-T-related toxicity.